The discovery of endothelin-1 (ET-1), a 21-amino acid peptide, has helped improve knowledge of local regulation of vascular tone by blood vessels (M. Yanagisawa et al. (1988) Nature 332(6163):411-5). ET-1 is generated in endothelial cells and vascular smooth muscle cells via conversion of proET-1 to ET-1 in the presence of endothelin converting enzyme-1 (ECE-1). This conversion from proET-1 to ET-1 is essential for optimal vasoconstrictor activity of ET-1 (M. Yanagisawa et al. (1988) Nature 332(6163):411-5 and G. D. Johnson et al. (1999) J Biol Chem 274(7):4053-8).
ET-1 is released from cultured endothelial cells at a slow basal rate. Due to a high vasoconstrictor potency and long lasting action, the continuous release of small amounts of ET-1 from endothelial cells towards the underlying smooth muscle cells may contribute to the maintenance of vascular tone and blood pressure (T. Miyauchi et al. (1999) Annu Rev Physiol 61:391-415). Under physiological conditions, the basal tone maintained by ET-1 is balanced by the release of endothelium derived relaxing factor (EDRF or nitric oxide and prostacyclin) and vasoconstrictor substances (thromboxane) (E. L. Schiffrin (1994) Clin Invest Med 17(6):602-20 and P. B. Persson (1996) Physiol Rev 76(1):193-244).
ET and its axis (ET-1, ET-2, ET-3, ETA and ETB receptors) have triggered considerable efforts to develop ET receptor antagonists having therapeutic potential in treating diseases like hypertension, heart failure, renal diseases, and cancer (A. Gulati et al. (1992) Drug Develop Res 26:361-387; A. Gulati et al. (1997) Neuropeptides 31(4) 301-9; G. Remuzzi et al. (2002) Nat Rev Drug Discov 1(12):986-1001; J. Nelson et al. (2003) Nat Rev Cancer 3(2):110-6; and A. Gulati et al. (2004) J Cardiovasc Pharmacol 44:S483-S486). Several ETA receptor antagonists, e.g., atrasentan, avosentan, clazosentan, darusentan, sitaxsentan, and ZD4054, are in mid to late stage clinical trial. Bosentan, a non-specific ETA and ETB receptor antagonist, has been marketed for a few years, and ambrisentan (ETA receptor antagonist) recently was approved for sale by the U.S. Food and Drug Administration (FDA) for a once-daily treatment of pulmonary arterial hypertension.
Intense efforts are devoted to develop ETA receptor antagonists. However, virtually no effort has been expended to develop ET agonists as therapeutic agents. The first proposed therapeutic use of an ETB receptor agonist resulted from a discovery that IRL-1620, a potent ETB receptor agonist, selectively enhanced breast tumor perfusion in rats (A. Rai et al. (2003) Cancer Chemother Pharmacol 51(1):21-8; A. Gulati (2003) U.S. Patent Publication 2004/0138121; and A. Gulati (2006) U.S. Patent Publication 2006/0211617). Administration of BQ788, a highly selective ETB receptor antagonist, blocked the tumor perfusion induced by IRL-1620 and confirmed the involvement of ETB receptors in tumor vasodilation (A. Rai et al. (2005) J Pharm Pharmacol 57(7):869-76 and N. V. Rajeshkumar et al. (2005) Breast Cancer Research and Treatment 94(3):237-247). The selective enhancement of tumor blood flow resulted in a greater percentage of infused paclitaxel reaching the tumor as compared to the normal tissues.
In a study conducted in breast tumor rats, IRL-1620 administration prior to paclitaxel resulted in a significant reduction of tumor volume, as well as a 20% complete remission of tumors, compared to paclitaxel treated rats (A. Rai et al. (2005) J Pharm Pharmacol 57(7):869-76. and N. V. Rajeshkumar et al. (2005) Breast Cancer Research and Treatment 94(3):237-247). See United States Patent Publication Nos. 2004/0138121, 2006/0211617, 2006/0257362, and 2007/0032422.
The present invention is directed to a new use for ETB receptor agonists, including IRL-1620, in the treatment of stroke and other cerebrovascular accidents. In particular, it now has been found that an ETB receptor agonist significantly increases cerebral blood perfusion, which is a novel and unexpected finding.
ETs are widely distributed throughout the body and are involved in a variety of physiological functions (A. Gulati et al. (1992) Drug Develop Res 26:361-387 and J. Nelson et al. (2003) Nat Rev Cancer 3(2):110-6). ETs exert their effects by binding to two distinct types of cell surface receptors, ETA and ETB ETA receptors have equal affinity for ET-1 and ET-2, and low affinity for ET-3. ETB receptors have equal affinity for ET-1, ET-2, and ET-3. Pharmacological evidence suggests that ETB receptors can be divided into two subtypes, i.e., ETB1 receptors present on endothelial cells and ETB2 receptors present on smooth muscle cells (D. P. Brooks et al. (1995) J Cardiovasc Pharmacol 26 Suppl 3:S322-5 and A. Leite-Moreira et al. (2004) Am J Physiol Heart Circ Physiol 287(3):H1194-9). Both ETA and ETB receptors belong to the G protein-coupled receptor (GPCR) family (J. Nelson et al. (2003) Nat Rev Cancer 3(2):110-6). ETA and ETB receptors located on vascular smooth muscle cells, produce vasoconstriction, whereas ETB receptors present on endothelial cells are mainly vasodilatory (G. Remuzzi et al. (2002) Nat Rev Drug Discov 1(12):986-1001).
IRL-1620 (N-Succinyl-[Glu9, Ala11,15] Endothelin 1) is a synthetic analogue of ET-1, i.e., a fragment of ET-1 having amino acids 8-21 of ET-1. IRL-1620 is a highly selective endothelin B receptor agonist, being 120,000 times more selective to ETB receptors than to ETA receptors (M. Takai et al. (1992) Biochem Biophys Res Commun 184(2):953-9). IRL-1620 has a molecular formula of C86H117N17O27 and a molecular weight of 1820.95. The molecular structure of IRL-1620, as illustrated in FIG. 1, is an amino acid sequence of Suc-Asp-Glu-Glu-Ala-Val-Tyr-Phe-Ala-His-Leu-Asp-Ile-Ile-Trp (SEQ ID NO:1).
Pharmacological Effects of IRL-1620
IRL-1620, like endothelins, can produce both vasodilation and vasoconstriction. Interaction of IRL-1620 with ETB receptors on endothelial cells leads to vasodilation, whereas an interaction with ETB receptors on smooth muscle cells leads to vasoconstriction. Furthermore, primary activation of ETB receptors by IRL-1620 can lead to autocrine/paracrine ET-1 release that subsequently activates both ETA and ETB receptors (S. Noguchi, et al. (1996) Br J Pharmacol 118(6):1397-402). Thus, the net effect of IRL-1620 is related to a number of factors, including the type of tissue, the species, and the physiological conditions. There have been a number of studies on pharmacological effects of IRL-1620 because it is a highly selective agonist of ETB receptors and often is used to delineate the role of ETB receptors in a given physiological situation. Some of these studies summarized below show that the vasoconstrictive effects of IRL-1620 are much less pronounced than those of ET-1. Other ETB receptor agonists known to persons skilled in the art produce pharmacological effects similar to those of IRL-1620, with the net effect also being related to the ability of a specific compound to selectively agonize ETB receptors.
Systemic Hemodynamic Effects
IRL-1620 exhibits systemic hemodynamic effects, including transient vasodilation and sustained vasoconstriction, in anesthetized rats (B. Palacios et al. (1997) Br Pharmacol 122(6):993-8 and S. W. Leung et al. (2002) J Cardiovasc Pharmacol 39(4):533-43), in an open-chest rat model (M. E. Beyer et al. (1995) J Cardiovasc Pharmacol 26 Suppl 3:S150-2), and in normal and cardiomyopathic hamsters (J. C. Honore et al. (2002) Clin Sci (Lond) 103 Suppl 48:280 S-283S). The vasoconstrictive effects of IRL-1620 are less pronounced compared to those of ET-1 (Palacios et al. (1997) Br J Pharmacol 122(6):993-8; J. C. Honore et al. (2002) Clin Sci (Lond) 103 Suppl 48:280 S-283S; and S. W. Leung et al. (2002) J Cardiovasc Pharmacol 39(4):533-43) and IRL-1620 had a positive inotropic effect (M. E. Beyer et al. (1995) J Cardiovasc Pharmacol 26 Suppl 3:S150-2).
Regional Hemodynamic Effects
IRL-1620 causes renal vasodilation in anesthetized dogs upon intrarenal arterial perfusion (T. Yukimura et al. (1994) Eur J Pharmacol 264(3):399-405) and pulmonary vasodilation in neonatal lambs upon intrapulmonary arterial injection (J. Wong et al. (1995) J Cardiovasc Pharmacol 25(2):207-15). A pulmonary vasodilatory effect of IRL-1620 also is observed in isolated perfused rat lungs (M. Muramatsu et al. (1999) Am J Physiol 276(2 Pt 1):L358-64). Injection of IRL-1620 into the circumflex coronary artery of anesthetized goats does not cause coronary vasoconstriction, whereas ET-1 administered similarly caused coronary vasoconstriction (J. L. Garcia et al. (1996) Eur J Pharmacol 315(2):179-86).
Effect on Respiratory Airway Smooth Muscles
Intravenous administration of IRL-1620 to anesthetized, artificially-ventilated guinea pigs resulted in bronchoconstriction in a biphasic manner (S. Noguchi et al. (1996) Br. J Pharmacol 118(6):1397-402). The second phase of bronchoconstriction probably is due to the activation of ETB receptors by IRL-1620 leading to autocrine/paracrine release of ET-1 that subsequently activated both ETA and ETB receptors (S. Noguchi et al. (1996) Br J Pharmacol 118(6):1397-402).
Experimental Studies on Human Tissues
In vitro, IRL-1620 causes contraction of human internal mammary arterial segments, but not human radial arterial segments (J. J. Liu et al. (1996) Clin Sci (Lond) 90(2):91-6). The contractile effect of IRL-1620 on internal mammary arteries reached a maximum of 20% of that obtained with ET-1 or noradrenaline. Further increases in concentration of IRL-1620 caused relaxation of the contracted arteries. IRL-1620 also had a contractile effect on human bronchial rings in a biphasic manner (T. Takahashi et al. (1997) Eur J Pharmacol 324(2-3):219-22).
Clinical Studies
To date, IRL-1620 has not been administered to humans. However, a phase I, open label, ascending dose study of the safety, tolerability, pharmacokinetics, and pharmacodynamics of IRL-1620 in patients with recurrent or progressive carcinoma (NCT00613691) is ongoing. Furthermore, a number of human studies have been conducted with ET-1, a much more potent vasoconstrictive agent than IRL-1620, as demonstrated in animal studies (B. Palacios et al. (1997) Br J Pharmacol 122(6):993-8 and S. W. Leung et al. (2002) J Cardiovasc Pharmacol 39(4):533-43). Administration of ET-1 to human subjects by perfusion at doses ranging from 1 to 20 ng/kg/min caused dose-dependent systemic vasoconstriction and consequential changes in hemodynamic parameters (D. Kiely et al. (1997) Cardiovasc Res 33(2):378-86; A. Franco-Cereceda et al. (1999) Scand Cardiovasc J 33(3):151-6; and F. Kiefer et al. (2000) Exp Clin Endocrinol Diabetes 108(5):378-81), but did not produce any serious adverse events.
Intravenous administration of ET-1 also causes coronary vasoconstriction (J. Pernow et al. (1996) Circulation 94(9):2077-82). However, coronary vasoconstriction may not be expected with IRL-1620 in humans. It has been shown that, in human coronary arteries, ETB receptors are absent or present at very low levels, and therefore, would make minimal contribution toward coronary vasoconstriction (W. A. Bax et al. (1994) Br J Pharmacol 113(4):1471-9; A. P. Davenport et al. (1995) J Cardiovasc Pharmacol 26 Suppl 3:S265-7; A. P. Davenport et al. (1994) Br J Pharmacol 111(1):4-6; W. A. Bax et al. (1993) Naunyn Schmiedebergs Arch Pharmacol 348(4):403-10; A. P. Davenport et al. (1995) J Cardiovasc Pharmacol 22 Suppl 8:522-5; and O. Saetrum Opgaard et al. (1996) Regul Pept 63(2-3):149-56).
Human studies also were conducted with an endothelin agonist, sarafotoxin S6c, which is less selective for ETB receptors than IRL-1620. On infusion into brachial artery, sarafotoxin S6c showed less reduction in forearm blood flow compared to ET-1 (W. G Haynes et al. (1995) Circulation 92(3): 357-63). Thus, any vasoconstrictive effects of IRL-1620 in humans are expected to be less than those observed with ET-1 and other endothelin agonists administered to humans to date.
Effect on Cerebral Blood Vessels
Endothelin has been implicated in a number of cerebrovascular disorders, including subarachnoid hemorrhage (R. Suzuki et al. (1992) J Neurosurg 77(1):96-100) and ischemic stroke (I. Ziv et al. (1992) Stroke 23(7):1014-6). It has been found that ETA receptor antagonists relieve chronic cerebral vasospasm (M. Clozel et al. (1993) Life Sci 52(9):825-34; S. Itoh et al. (1993) Biochem Biophys Res Commun 195(2):969-75; H. Nirei et al. (1993) Life Sci 52(23):1869-74; and R. N. Willette et al. (1994) Stroke 25(12):2450-5; discussion 2456). Studies have been performed to characterize endothelin receptors in the cerebral blood vessels. ETA receptors were found to mediate contraction in human cerebral, meningeal, and temporal arteries (M. Adner et al. (1994) J Auton Nerv Syst 49 Suppl:S117-24 and a marked ETB receptor-mediated relaxation was obtained with ET-3 when ETA receptor activity was blocked using FR139317 (IUPAC Name: (2R)-2-[[(2R)-2-[[(2S)-2-(azepane-1-carbonylamino)-4-methylpentanoyl]amino]-3-(1-methylindol-3-yl)propanoyl]amino]-3-pyridin-2-ylpropanoic acid) in precontracted human temporal arteries (G. A. Lucas, et al. (1996). Peptides 17(7): 1139-44).
Overall, a need still exists in the art to identify agents, or combinations of agents, that effectively treat strokes and other cerebrovascular accidents. To date, no report exists on the effect of IRL-1620 on cerebral circulation, and the present disclosure is the first reporting that IRL-1620 increases cerebral blood perfusion, as measured with laser-Doppler perfusion method.